Small molecule modulation of the p75 neurotrophin receptor suppresses age- and genotype-associated neurodegeneration in HIV gp120 transgenic mice

Highlights

• HIV gp120 and aging synergize to accelerate inflammation and neurodegeneration.

• Extracellular Tau and p75 receptor deposits co-accumulate in the aging hippocampus.

• Inflammation and neurodegeneration are suppressed by a novel p75 receptor ligand.

• Hippocampal inhibitory interneurons are preserved by the p75 receptor ligand.

• Modulation of p75 has significant therapeutic potential in people living with HIV.

Abstract

The persistence of HIV in the central nervous system leads to cognitive deficits in up to 50% of people living with HIV even with systemic suppression by antiretroviral treatment. The interaction of chronic inflammation with age-associated degeneration places these individuals at increased risk of accelerated aging and other neurodegenerative diseases and no treatments are available that effectively halt these processes. The adverse effects of aging and inflammation may be mediated, in part, by an increase in the expression of the p75 neurotrophin receptor (p75^NTR) which shifts the balance of neurotrophin signaling toward less protective pathways. To determine if modulation of p75^NTR could modify the disease process, we treated HIV gp120 transgenic mice with a small molecule ligand designed to engage p75^NTR and downregulate degenerative signaling. Daily treatment with 50 mg/kg LM11A-31 for 4 months suppressed age- and genotype-dependent activation of microglia, increased microtubule associated protein-2 (MAP-2), reduced dendritic varicosities and slowed the loss of parvalbumin immunoreactive neurons in the hippocampus. An age related accumulation of microtubule associated protein Tau was identified in the hippocampus in extracellular clusters that co-expressed p75^NTR suggesting a link between Tau and p75^NTR. Although the significance of the relationship between p75^NTR and Tau is unclear, a decrease in Tau-1 immunoreactivity as gp120 mice entered old age (>16 months) suggests that the Tau may transition to more pathological modifications; a process blocked by LM11A-31. Overall, the effects of LM11A-31 are consistent with strong neuroprotective and anti-inflammatory actions that have significant therapeutic potential.

Introduction

HIV rapidly enters the nervous system resulting in a chronic infection, inflammatory responses and progressive neural damage. The neural damage has been linked, in part, to the release of soluble factors from microglia (MG) and macrophages (M) (Giulian et al., 1990; Pulliam et al., 1991; Bragg et al., 2002; Kolson, 2002; Meeker et al., 2012) as well as direct neural effects of HIV proteins. These factors lead to cognitive dysfunction in up to half of the individuals infected with HIV. Although antiretroviral therapy (ART) has reduced the severity of cognitive dysfunction through effective long-term control of systemic virus, HIV persists in the CNS and cognitive decline continues. As HIV infected individuals maintained on ART approach advanced ages, they are also displaying an accelerated aging-like phenotype that may synergize with other age-associated diseases such as Alzheimer's disease (AD) (Martin et al., 2013; Harper, 2015; Gianesin et al., 2016; Levine et al., 2016; Cohen and Torres, 2017; Pfefferbaum et al., 2018). Synergy between HIV infection and aging may be mediated in part through the effects of inflammation on neurons which are similar in both disease processes(Stokin et al., 2005; Kuchibhotla et al., 2008; Bittner et al., 2010; Meeker et al., 2016a). The potential for synergy is highlighted in postmortem studies that have identified aggregates of amyloid beta (Aβ) in HIV infected patients although the direct role of HIV is still debated(Fields et al., 2018; Stern et al., 2018; Fulop et al., 2019).

Efforts to reduce inflammation and neural damage have focused on ways to protect neurons or block the destructive inflammatory processes. Studies employing in vitro models of HIV induced inflammation and neuropathogenesis have shown that modulators that enhance neurotrophin signaling may offer strong neuroprotection. Alterations in nerve growth factor (NGF) signaling via TrkA and brain-derived neurotrophic factor (BDNF) signaling via TrkB are thought to contribute to both HIV- and age-associated neural dysfunction(Fahnestock et al., 2001; Peruzzi et al., 2002; Costantini et al., 2005; Nosheny et al., 2005; Cuello et al., 2007; Mufson et al., 2007; Nosheny et al., 2007; Capsoni et al., 2011; Liu et al., 2018). Neurotrophins and related ligands are protective in various rodent models of neuropathogenesis (Ebadi et al., 1997; Mocchetti and Bachis, 2004a; Longo et al., 2007; Mocchetti et al., 2007; Yang et al., 2008; Aloe et al., 2012; Capsoni et al., 2012; Knowles et al., 2013; Longo and Massa, 2013; Shi et al., 2013; Simmons et al., 2014) and in natural aging (Xie et al., 2019). Thus, modulation of neurotrophin signaling has substantial potential for the inhibition of both age- and disease-related cognitive decline.

The p75 neurotrophin receptor (p75^NTR) has proven to be an attractive target for modification of neurotrophin signaling(Longo and Massa, 2013; Meeker and Williams, 2014). In the context of aging and AD, an increase in the expression of the p75^NTR along with decreased levels of NGF and increased levels of proNGF are thought to contribute to the increased vulnerability of the aging nervous system by shifting neurotrophin signaling away from beneficial NGF/TrkA signaling toward deleterious proNGF/p75^NTR signaling(Fahnestock et al., 2001; Counts et al., 2004; Costantini et al., 2005; Counts and Mufson, 2005; Ginsberg et al., 2006; Mufson et al., 2007; Capsoni et al., 2010; Capsoni et al., 2011; Cuello et al., 2012; Mufson et al., 2019).

A novel ligand, LM11A-31, has been developed that binds p75^NTR to down regulate degenerative signaling, upregulate survival signaling and prevent deficits associated with pro-neurotrophin signaling (Massa et al., 2006). The compound is orally bioavailable, crosses the blood brain barrier and has no known side effects at therapeutic concentrations. Using in vitro models of HIV neuropathogenesis, we have shown that low nanomolar concentrations of LM11A-31 provide neuroprotection against the effects of virus and the viral envelope protein, gp120(Meeker et al., 2012; Meeker et al., 2016a). Similar protective effects have been seen in a variety of in vitro and in vivo disease models including AD (Yang et al., 2008; Knowles et al., 2013; Simmons et al., 2014) Huntington disease (Simmons et al., 2016), traumatic brain injury (Shi et al., 2013) and models involving pathological accumulations of proNGF including spinal cord injury (Tep et al., 2013), bladder degeneration (Ryu et al., 2018) and juvenile indiopathic arthritis (Minnone et al., 2017).

In addition to direct neuroprotection, LM11A-31 may restrict inflammation. Studies of cultured human monocyte-derived macrophages (hMDM) have shown expression of TrkA and the p75^NTR which respond to their respective ligands, mature NGF and its precursor, proNGF, to control the phenotype of these cells. NGF was shown to induce a morphological and functional phenotype the opposite of proNGF suggesting a prominent role for neurotrophin signaling in the control of macrophage activity (Williams et al., 2015). Macrophages stimulated with proNGF displayed a phenotype that enhanced damage to neurons whereas NGF was partially protective. This dichotomy in the actions of NGF versus proNGF is similar to the proposed actions on neurons where a shift in the neurotrophin balance to higher proNGF:NGF ratios predisposes neurons to degeneration during aging and Alzheimer's disease(Fahnestock et al., 2001; Peng et al., 2004; Al-Shawi et al., 2008; Zeng et al., 2011; Mufson et al., 2019). These studies suggest that neurotrophin ligands may have beneficial anti-inflammatory effects on macrophages and microglia. This may be particularly true for a p75^NTR ligand given the role of proNGF/p75^NTR signaling in the activation of neurotoxic responses in macrophages.

The robust neuroprotective effects combined with potential anti-inflammatory effects suggest that LM11A-31 may have substantial therapeutic benefit. In the current study, we used the gp120 transgenic mouse engineered to express the HIV envelope protein gp120_IIIB to examine the ability of LM11A-31 to modify the development of inflammation and neural damage. The model recapitulates many of the features of HIV infection including the gradual development of inflammation making it a good model to investigate the interactions between HIV-associated inflammation and aging. LM11A-31 was found to suppress inflammation and neural damage in support of a potential therapeutic effect in HIV patients.